The present invention relates to transgenic animals. Specifically, the invention relates to transgenic fish and methods for their use.
Animal models of disease states play an important role in identifying the underlying biochemical mechanisms of particular diseases, as well as discovering therapeutic agents to eradicate the disease or otherwise lessen its symptoms. For example, rabbit models of familial hypercholesterolemia, rat models of non-insulin-dependent diabetes mellitus, mouse models of cancer and hamster models for spontaneous atrial thrombosis are known. Additionally, animal models for genetic diseases have arisen spontaneously in a variety of species, including mice, cats and dogs. Working with such large animals poses several drawbacks.
For example, many of the animals used in such models are relatively large vertebrates which take up a large amount of research space, are costly to feed and otherwise maintain, have slow reproductive cycles, produce relatively few offspring at one time, and cannot effectively mimic all desired disease states. Researchers have attempted to obtain animal models that solve some of these problems, but have not yet obtained such animal models for all desired diseases. For example, transgenic fish models of premalignant and non-neoplastic or non-malignant hyperproliferative disorders (e.g., inflammation and retinopathy) would also be useful. Additionally, although fish have been utilized to detect mutagens in aquatic environments, there are currently no transgenic fish models that develop any cancers relevant in human cancer research, including, for example, human T-cell leukemias, non-Hodgkin's lymphoma, high-grade astrocytoma, rhabdomyosarcoma, neuroblastoma, neuorendocrine carcinoma, pancreatic carcinoma, ovarian carcinoma, testicular carcinoma, stomach cancer, colon cancer, renal cancer, melanoma and acute or chronic myeloid leukemia.
Human T-cell leukemias can arise from oncogenes activated by specific chromosomal translocations involving T-cell receptor genes. In particular, T-cell acute lymphoblastic leukemia is a malignant disease of thymocytes, accounting for about 10% to about 15% of pediatric and about 25% of adult acute lymphoblastic leukemia cases. Although some therapies are available for T-cell acute lymphoblastic leukemia, they are not as effective as desired.
Follicular center cell non-Hodgkin's lymphomas are a common and generally indolent type of B-cell lymphoma that occurs almost exclusively in adults, and 80% of follicular center cell lymphomas have a t(14;18) chromosomal translocation. Molecular analysis of the breakpoints of the 14;18 translocation identified BCL2 as the gene on chromosome 18 that is overexpressed due to its translocation into the IgH locus on chromosome 14. Functional studies revealed that BCL2 defines a new class of proto-oncogene products that act to prolong cell survival, rather than through more typical effects on cell differentiation or proliferation. It has since been learned that BCL2 is a member of a large family of highly conserved proteins that either inhibit or promote apoptosis. In addition, BCL2 and it's pro-survival relatives may be important for the aberrant survival of many human cancers, not just those with overexpression of BCL2 due to the t(14;18).
High-grade astrocytomas are among the most common and devastating adult brain tumors, spreading so rapidly that patients seldom survive more than 9-12 months. Despite progress in surgical, radiation and chemotherapy technologies, there has been little improvement in the outcome of patients with astrocytoma over the last twenty years. Clearly, novel approaches are needed to better understand the biological basis of this disease before effective therapies can be developed.
Rhabdomyosarcomas are a heterogeneous group of malignant tumors of skeletal muscle progenitors and are the most common soft-tissue sarcoma in children of 15 years or younger. Rhabdomyosarcoma consists of two histologic subtypes, alveolar and embryonal, each characterized by the misexpression of different subsets of genes. The aggressive nature of these tumors makes their effective treatment particularly difficult. While rhabdomyosarcomas can be observed in genetically engineered, mammalian disease models, they are often associated with other tumor types. While informative, a more specific model of rhabdomyosarcoma is necessary to elucidate its molecular basis and to identify novel genes that may ultimately be used as targets for the development of novel therapeutic strategies.
The mutations and gene rearrangements commonly seen in acute myeloid leukemias typically result from a chromosomal translocations such as the t(8;21) or t(15;17), generate chimeric oncoproteins by fusing one or two transcription factors (Look, 1997). However, these alterations are not sufficient to explain the induction of acute leukemia. Additional animal models are needed, for example, to permit the unbiased detection of mutations in many potentially novel genes that lead to leukemia, which is currently not possible in other mammalian models.
Many of the underlying mechanisms that lead to neuroblastoma, neuorendocrine carcinoma, pancreatic carcinoma, ovarian carcinoma, testicular carcinoma, stomach cancer, colon cancer, renal cancer, melanoma and acute or chronic myeloid leukemia have yet to be fully understood. Identifying the genes mutated in these diseases will lead to new insights into cancer as a whole. Additionally, using a vertebrate model system in which genetic or chemical suppressors can be identified that inhibit or delay disease progression, or sensitivity to chemotherapy or radiation-induced programmed cell death, will be necessary to identify new drug targets for the development of targeted chemotherapies. For example, a model system is needed, which does not require an a priori knowledge of the specific target. Target elucidation may be accomplished after the modulating target drug or agent is demonstrated safe and effective, which, thus, saves both time and expense in the drug discovery process.
A further understanding of the cellular and molecular genetic features of various disease states such as the cancers listed above are needed. An appropriate animal model would be invaluable to elucidate the multistep process of genetic mutations, as well as to develop more effective drugs. The present invention addresses these needs.